Genetic diversity among a complex of cereal cyst nematodes inferred from RFLP analysis of the ribosomal internal transcribed spacer region

This study examined the restriction polymorphism (RFLP) of the nuclear ribosomal DNA in Heterodera avenae, H. filipjevi, H. mani, H. latipons, and the taxonomically unclear Gotland strain in order to establish a molecular characterization and phylogenetic relationships in the complex of cereal cyst nematodes (CCN). The internal transcribed spacer (ITS) and 5.8S rDNA were amplified by PCR from a single female or a cyst of 27 different geographic isolates of the CCN complex and one population of H. schachtii, used as outgroup. The amplified product was 1.2 kb long and 14 of 15 enzymes produced restriction fragments for each isolate. Relationships between populations were determined from UPGMA analysis based on distance values calculated from RFLP data. Digestions with TaqI clearly differentiated H. avenae, H. latipons, and a group composed of H.filipjevi and the Gotland strain. Six endonucleases (HaeIII, HinfI, ItaI, PstI, TaqI, and Tru9I) produced the same restriction pattern with H. filipjevi and the Gotland strain, and both were clearly separated from H. avenae with PstI. Restriction sites have revealed a mixture of the species H. latipons and H. avenae, and possible infraspecific variation in H. avenae. The inferred phylogenetic relationships of species in the CCN complex are in agreement with their morphological characterization.